磁珠富集法筛选实验豚鼠微卫星分子标记

Microsatellite Enrichment by Magnetic Beads in Guinea Pig (Cavia porcellus)

目的直接从实验豚鼠基因组DNA中筛选获得微卫星分子标记。方法应用磁珠和生物素标记的微卫星探针与豚鼠基因组酶切片段杂交,捕获200～1000 bp含有微卫星序列的DNA片段,连接到pMD-18V载体中,转化到感受态细胞E.coli DH5α中构建富集微卫星序列的小片段插入文库。然后用PCR法进行筛选。结果从约2000个转化子中获得240个阳性克隆。对其中98个进行了测序,并成功设计豚鼠微卫星引物17对。结论经过优化的磁珠富集法能够稳定、高效地获得豚鼠微卫星标记。本研究获得的微卫星位点将成为豚鼠遗传学研究的有力工具。

Objective To isolate microsatellite loci from guinea pig（Cavia porcellus） genome.Methods The 200 ～ 1000 bp DNA fractions of guniea pig containing microsatellite sequences were captured by hybridization with the oligonucleotide probes attached to streptavadin coated magnetic beads.The enriched DNA were ligated into pMD-18T and then transformed into E.coli DH5α competent cells.The method of PCR was used to screen positive clones from the libraries.And microsatellite primers were disigned based on the sequences from the positive clones.Results In total 240 positive clones were identified from about 2000 transformants in the libraries screened by PCR.Finally,we got 98 sequences and successfully designed 17 pairs of microsatellite primers for guniea pig.Conclusions The results showed that this method is very efficient to isolate microsatellite markers and the markers are useful tools for further studies on guinea pig genetics.